Efficacy and safety of oral alitretinoin versus oral azathioprine in patients with severe chronic hand eczema: Results from a prematurely discontinued randomized controlled trial

Alitretinoin is the only registered systemic treatment of severe chronic hand eczema (HE), but it is most effective in hyperkeratotic HE. It has never been compared in head-to-head studies to other immunomodulating systemic drugs, for example, azathioprine, that can be used (off-label) for severe HE. In one daily practice case series, patients showed moderate-togood improvement of their HE after treatment with azathioprine. Based on this case series, in which only one patient with hyperkeratotic HE was included, combined with observations in clinical practice, azathioprine could prove superior to alitretinoin in HE subtypes other than hyperkeratotic HE. This trial aimed to compare alitretinoin to azathioprine in the treatment of severe chronic HE. Due to a high-drop out ratio, the trial was unfortunately prematurely discontinued. Most prematurely discontinued studies remain unpublished, resulting in unwanted publication bias. Therefore, we decided to analyse our data.

(QALYs) and cost-effectiveness will be assessed with the EQ-5D-5L questionnaire while monitoring treatment related costs.
Nature and extent of the burden and risks associated with participation, benefit and group relatedness: this trial is designed in a way that minimizes the burden and risks for the patient, because it will be carried out according to daily practice at our department. No additional laboratory measurements will be carried out next to measurements that are routinely assessed in treatment with one of both study drugs. One extra visit is needed to give patients consideration time before participation in the study. Results of the trial can be related to the population of patients with severe chronic non-hyperkeratotic hand eczema.

INTRODUCTION AND RATIONALE
Hand eczema is a common skin disease of the hands. 50% of patients reported a beneficial effect of the treatment. (13) In an Indian randomized comparative trial, conducted in 2013, chronic hand eczema patients were treated with either topical clobetasol propionate 0,05% cream alone, or in combination with azathioprine. A 91% improvement of hand eczema severity index (HECSI) score at 24 weeks was found in the azathioprine group, compared to 39% in the control group. (14) In patients with atopic dermatitis (AD) who were treated with azathioprine a mean relative reduction in severity scoring of atopic dermatitis (SCORAD) was found in 39% of Although these studies show favorable results, more studies on the use of azathioprine in the treatment of hand eczema are needed. This trial aims to compare alitretinoine to azathioprine in the treatment of severe chronic non-hyperkeratotic hand eczema. The study assesses the efficacy of both treatments and will show head-to-head results, which should contribute to uncovering the best treatment strategy for hand eczema.

OBJECTIVES
Primary objective: to compare the efficacy of alitretinoine and azathioprine in the treatment of patients with severe chronic non-hyperkeratotic hand eczema.
Secondary objectives: • to compare time to response • to compare health related quality of life • to compare improvement in severity of hand eczema, assessed by the patient • to compare safety • to compare cost-utility and cost-effectiveness

STUDY DESIGN
This study is designed as a randomized prospective open label study (or RCT). It will be conducted at the Department of Dermatology of the University Medical Center of Groningen, a tertiary referral center. Assessment of disease severity, laboratory measurements and quality of life in this study will be conducted comparable to daily practice assessments at our department.
The duration of the study for an individual patient is 24 weeks. The expected total duration of the study is 1.5 years.
Flow chart of study design:

Population (base)
The study population will exist of adult patients with severe chronic non-hyperkeratotic hand eczema, visiting the Department of Dermatology of the University Medical Center Groningen.
Non-hyperkeratotic hand eczema will be diagnosed following the criteria of the Danish Contact Dermatitis Group. (6) The severity of the hand eczema at the screening will be graded by means of a Physician Global Assessment using a validated Photoguide. (16) Woman in the fertile age will be required to use proper contraception methods. Men and women of all ethnicities of 18 years and older will be recruited.
Patients meeting all inclusion criteria, while not meeting any of the exclusion criteria, will be asked to participate.
Our sample size calculation showed a total of 116 patients need to be included. Given the fact that 300-350 individual patients with hand eczema visit our outpatient eczema clinic every year, and the mean severity is high because our department is a tertiary referral center, we believe this number to be feasible.

Inclusion criteria
In order to be eligible to participate in this study, a subject must meet all of the following criteria: • Age ≥ 18 years and ≤ 75 years • Severe chronic non-hyperkeratotic hand eczema for a minimum duration of 3 months as defined by a Physician Global Assessment (PGA) using a validated Photoguide (16) • Refractory to standard therapy, defined as:

Exclusion criteria
A potential subject who meets any of the following criteria will be excluded from participation in this study: General criteria prior to randomization • Treatment with alitretinoin or azathioprine in the previous 3 months • Hyperkeratotic palmar eczema as defined by the Danish Contact Dermatitis Group (6) • Patients with predominantly atopic dermatitis, in which the hands are also involved.
Patients with mild atopic dermatitis, in which the hands are mainly affected are eligible for inclusion.

Sample size calculation
This trial hypothesizes a superior response to azathioprine compared to alitretinoin in the treatment of severe chronic non-hyperkeratotic hand eczema.
A sample size of 52 in each group will have 80% power to be able to reject the null hypothesis of no difference between alitretinoin and azathioprine, using a chi-square test with a two-sided 0.05 significance level. In this calculation we used the following assumptions: randomization ratio is 1:1, and we expected the percentage of responders in the alitretinoin group to be 33% (10). Literature on the efficacy of azathioprine on hand eczema in terms of PGA is lacking. From a retrospective assessment of available patient data from our department, combined with clinical experience, we estimate 60% responders in the azathioprine group. We anticipate a drop-out of maximal 10% of randomized patients; a small percentage prior to first application of study drugs due to excluding laboratory measurements and a larger percentage during follow up, mainly due to subjective side effects. We therefore plan to include 116 patients in total, 58 in the alitretinoin group and 58 in the azathioprine group. (

Investigational product/treatment
Group I will receive alitretinoin.
Group II will receive azathioprine.
Dosage, dosage modification and method of administration can be found in paragraph 6.6.

Use of co-intervention
All patients will be given an emollient cream with instructions to apply it frequently. One week before the first intake of study drugs, concomitant treatment with a topical class II corticosteroid at maximum is permitted when needed, with a maximum application of one finger-tip-unit (FTU) for each hand daily. (17)  Azathioprine specific prohibited concomitant treatment: allopurinol, febuxostat, ribavirin.
Patients on cumarines are requested to inform their thrombosis service of treatment with azathioprine. Careful monitoring of coagulation is advised. (18)

Escape medication
In case of an exacerbation or postponed treatment effect, patients are allowed to receive a maximum of 3 courses of rescue medication: mometasone furoate once daily for 1 week, with a maximum application of one FTU for each hand daily. (17)

INVESTIGATIONAL PRODUCT
The products of this study will be used as in usual clinical practice.

Name and description of investigational products
Alitretinoin is a retinoid drug. The pharmacological action of retinoid drugs may be explained by their effects on cell proliferation, cell differentiation, apoptosis, angiogenesis, keratinization, sebum secretion and immunomodulation. Unlike other retinoids, which are specific agonists of either RAR or RXR receptors, alitretinoin binds to members of both receptor families. The mechanism of action of alitretinoin in chronic hand eczema is unknown. It has been shown that alitretinoin has immunomodulatory and antiinflammatory effects that are relevant to skin inflammation. CXCR3 ligands and CCL20 chemokines, that are brought to expression in eczematous skin lesions, are downregulated by alitretinoin in cytokine stimulated keratinocytes and dermal endothelium cells.
In addition, alitretinoin suppresses the expansion of cytokine activated leukocyte subsets and antigen presenting cells. It has been observed that in humans alitretinoin only minimally affects sebum secretion. (10) Azathioprine is a prodrug of 6-mercaptopurine (

Summary of findings from non-clinical studies
Alitretinoin (from SPC text) (10) Acute toxicity As with other retinoids, the acute toxicity of alitretinoin was low in mice and rats. The LD50 after intraperitoneal administration was >4000 mg/kg after 24 hours and 1400 mg/kg after 10 days. The approximate LD50 after oral administration in rats was 3000 mg/kg.

Chronic toxicity
Protocol ID 52232 Alitretinoin vs azathioprine in severe non-hyperkeratotic hand eczema Alitretinoin was tested in long-term studies up to 9 months in dogs and 6 months in rats.
Signs of toxicity were dose-related and occurred at exposures similar to the human therapeutic exposure based on area-under-the-curve (AUC). Effects were characteristic for retinoids (consistent with hypervitaminosis A), and were generally spontaneously reversible.

Teratogenicity
Like other retinoids, alitretinoin has been shown to be teratogenic in vitro and in vivo.
Due to the teratogenic potential of alitretinoin, women of childbearing potential must adhere to strict pregnancy prevention measurers during and 1 month following alitretinoin therapy.

Fertility
Alitretinoin was tested in a study of fertility and early embryonic development in rats. No effects on male or female reproductive parameters were observed at the highest dose tested. However, systemic exposure in this study did not reach the level observed in patients.
As with other retinoids reversible effects on male reproductive organs were observed in experimental animals in the form of disturbed spermatogenesis and associated degenerative lesions of the testes. The safety margin in dogs with regard to the no-effect level of toxicity to male reproductive organs was 1-6 for a human dose of 30 mg.

Mutagenicity
In in vitro or in vivo tests, alitretinoin has been shown not to be mutagenic.

Carcinogenicity
Alitretinoin was tested in 2-year carcinogenicity studies in rats and mice. Dose-related retinoid-specific toxicity was seen at higher doses, but no carcinogenic potential was noted.

Phototoxicity
Alitretinoin was found to be phototoxic in vitro and in vivo Azathioprine (from SPC text) (18) Toxicological studies in animals have shown that the hematopoietic system is most strongly influenced by depression of mainly granulopoiesis and relative sparing of megakaryocytes and thus the formation of thrombocytes. In dogs azathioprine caused Teratogenicity or embryolethality has been seen in a number of animal species with varying degree of susceptibility. In rabbits, a dose of 5-15 mg/kg body weight daily on days 6-14 of pregnancy produced skeletal abnormalities, in mice and rats, doses of 1-2 mg/kg body weight daily on days 3-12 were lethal to embryos.
Evidence of teratogenicity of azathioprine in man is equivocal. As with all cytotoxic chemotherapy, contraceptive precautions should be advised when either partner is receiving azathioprine.
Azathioprine was mutagenic in a number of in-vitro and in-vivo genotoxicity assays.
In long-term carcinogenicity studies of azathioprine in mice and rats, an increased incidence of lymphosarcomas (mice) and epithelial tumours and carcinomas (rats) were observed at dosages that were up to 2-fold the human therapeutic dosage.

Summary of known and potential risks and benefits
The intended benefit of study drugs is to reduce the severity of hand eczema.
Main risks in the alitretinoin group are (10): -Teratogenicity of the study drugs -Reversible pneumonitis (rare)

Description and justification of route of administration and dosage
Group I will receive an oral alitretinoin capsule of 30mg once daily for a total of 24 weeks.
In a dose-finding study, the effectiveness and tolerability of this dose was established and it is recommended to use this as the standard dose for the prescription of alitretinoin in hand eczema. (8,10) Group II will receive oral azathioprine tablets twice daily in a dose of 1.5 or 2.5mg/kg/day, depending on the thiopurine methyltransferase (TPMT) activity. A blood sample to measure TPMT activity will be obtained at the randomization visit. In thiopurine metabolism, the activity of the gene that encodes for the expression of the TPMT enzyme has an important effect on the toxicity of thiopurines in the human body. Genetic interpersonal differences lead to a difference in TPMT activity. Patients with low or absent activity of TPMT (0.3% of patients) have a risk of fatal myelosuppression when given normal doses of thiopurines. The result of the test is available in 3-4 weeks in our center.
Patients will be allowed to continue their current topical treatment (if this is in accordance with the in-/exclusion criteria) for three more weeks and are required to taper the strength of topical corticosteroids to maximum class 2 during the week prior to baseline.
At the azathioprine baseline visit (4 weeks following randomization visit) patients receive a dosage of azathioprine tailored to their TPMT activity. Patient with a low or absent TPMT activity (indicating homozygous or compound heterozygous mutations) will be excluded from the study. Patients with intermediate activity of TPMT (indicating heterozygous mutation) will receive 1.5mg/kg/day. Dosage in patients with normal to high activity levels of TPMT (indicating homozygous wild-type) will receive 2.5mg/kg/day. This is in accordance with current guidelines. (20)

Dosages, dosage modifications and method of administration
The study drugs will be distributed to participants in amounts that allow drug intake as per protocol up to the next visit, 4 weeks later.
Dosage reduction is allowed in both groups in case of abnormal findings on physical examination, laboratory markers, and/or adverse events. Alitretinoin dose can be reduced from 30mg/day to 10mg/day, in accordance with the SPC-text.
Alitretinoin capsules need to be administered orally once daily, during a meal.
Azathioprine tablets need to be administered orally with a glass of liquid (200ml), during a meal.

Preparation and labelling of Investigational Medicinal Product
Preparation and labelling of the study drugs will be carried out according to usual practice by the community pharmacy, honouring relevant GMP guidelines.

Drug accountability
Not applicable.

NON-INVESTIGATIONAL PRODUCT
Not applicable. In this study the main endpoint is the between-group difference in response to treatment between baseline and 24 weeks of treatment.

Severity of hand eczema
• Between-group difference in mean change in PGA between baseline and 12 and 24 weeks.
• Between-group difference in response to treatment between baseline and 12 weeks of treatment.  Safety and tolerability • Adverse events in both groups will be registered.

Cost-utility and cost-effectiveness
• Between-group difference in mean Quality Adjusted Life Years (QALY's) will be measured by the EQ-5D-5L score at baseline, week 12 and week 24. The EQ-5D-5L is a measure for HRQoL and utility values. The EQ-5D-5L questionnaire includes a descriptive system, which comprises 5 dimensions of health: mobility, self-care, usual activities, pain/discomfort, and anxiety/depression.
Moreover it includes a visual analog scale (VAS), which records the respondent's self-rated health status on a graduated (0-100) scale.
• Direct medical costs will be determined using standardized prices for consultation, treatment (medication; alitretinoin or azathioprine, topical treatment of corticosteroids and emollientia; if necessary oral or topical treatment of antibiotics), diagnostic tests, laboratory measurements, visits to the general practitioner for hand eczema and hospital admissions (in patient and/or daycare). Included patients will be asked to keep track of how much they spend on over-the-counter medication and other products for their hand eczema (outof-pocket costs). Direct non-medical costs, consisting of travel costs, will be determined using average travel costs to the hospital as determined by relevant Dutch guidelines on cost-studies in healthcare. (22) • Indirect costs, consisting mainly of productivity loss, will be also be calculated using tables from the guidelines with average income of Dutch workers stratified by age and gender, corrected for shift working / irregular working hours.

Randomisation, blinding and treatment allocation
Randomization is carried out by a computer program. This is a study with blinded efficacy assessors, which are unaware of treatment allocation.
The participants and treating physician will be aware of treatment allocation. Efficacy assessment will be carried out by physicians or specialized eczema nurses who are experienced in assessing hand eczema by PGA and HECSI in daily pratice. Blinding will only be broken after analyzing the data.
At screening visit (-1 week for alitretinoin, -5 weeks for azathioprine), clinical eligibility of patients is assessed. If eligible, patients are given a least 1 week time to consider participation in the study. If patients choose to participate they will be randomized to alitretinoin or azathioprine treatment (at week 0 for alitretinoin, -4 weeks for azathioprine).
Based on the assigned group, appropriate laboratory assessments will be performed. If these assessments do not show abnormalities as described in the laboratory exclusion criteria post randomization, patients will receive the treatment.

Study procedures
Procedures part of standard medical treatment: According to daily practice, a detailed patient history is obtained of all newly referred patients with hand eczema, and they are planned for patch testing to exclude contact allergy. During this first period, patients are treated with topical corticosteroids and emollients. Standard skin care advice is provided and exposure to provoking factors is discussed. If a relevant contact allergy is ruled out and the hand eczema proves to be refractory to topical therapy and/or UV therapy, the next step is systemic therapy. These patients are a candidate for the current study.
Laboratory analysis is performed to verify contra-indications for systemic treatment; in the current study alitretinoin or azathioprine. For azathioprine this check takes 4 weeks, due to the necessary measurement of TPMT (see paragraph 6.5). During therapy, standard monitoring of blood values is carried out, according to SPC texts and current guidelines.
At every visit, the Physician Global Assessment (PGA) for severity is determined (9) and the hand eczema is scored using the Hand Eczema Severity Index (HECSI, see appendix) (21), corresponding to our daily practice. Additionally, a Patient Global Assessment (PaGA) for improvement is obtained at week 12 and 24. Furthermore, health related quality of life is scored with a Dutch version of the Quality of Life in Hand Eczema Questionnaire (QOLHEQ, see appendix) at the start of therapy, at week 12 and week 24.
Procedures extra for this study: Patients will be given one week to consider participation. Due to this, one extra visit is needed to randomize the patient and obtain baseline data. A Patient Global Assessment (PaGA) one-item questionnaire will be obtained at week 12 and 24. This procedure is only extra in terms of obtaining a quantitative assessment of the qualitative report that a patient provides us in daily practice.
Patients will be asked to keep track of out-of-pocket costs on products for their hand eczema. During each visit, patients will be asked for direct and indirect medical and nonmedical costs (see paragraph 8.1.2).
No diagnostic procedures or other treatments will be postponed for patients participating in this study. However, patients receive at least 1 week time to consider participation in the study.

Study overview:
In table 1 and 2 a systematic overview of the study for respectively the alitretinoin and azathioprine arm is presented.  Patients are permitted to deviate from the schedule with a maximum of 7 days during week 0-8. From week 9 a maximum deviation of 14 days is permitted.

Withdrawal of individual subjects
Subjects can leave the study at any time for any reason if they wish to do so without any consequences. The investigator can decide to withdraw a subject from the study for medical reasons.

Specific criteria for withdrawal
• Evidence of pregnancy

Replacement of individual subjects after withdrawal
Subjects will not be replaced after withdrawal.

Follow-up of subjects withdrawn from treatment
Subjects withdrawn from treatment will continue to be treated at our center, outside the study. All efforts will be made to report the observations for the reason(s) for premature withdrawal and the time of occurrence. If an adverse event is the reason for withdrawal, the physicians of our department will administer therapy as clinically indicated. All serious adverse events and those adverse events for which the relationship to the study drug is plausibly related will be followed up until they have returned to baseline status or stabilized.

Premature termination of the study
Alitretinoin is registered for the treatment of hand eczema. Azathioprine has been used for decades to treat hand eczema and atopic dermatitis, among other dermatoses. Therefore we do not expect a substantial number of unexpected serious adverse events.
If an unforeseen substantial amount of (unexpected) serious adverse events occur, we will consider premature termination of the study.

Section 10 WMO event
In accordance to section 10, subsection 1, of the WMO, the investigator will inform the subjects and the reviewing accredited METC if anything occurs, on the basis of which it appears that the disadvantages of participation may be significantly greater than was foreseen in the research proposal. The study will be suspended pending further review by the accredited METC, except insofar as suspension would jeopardise the subjects' health. The investigator will keep all subjects informed.

Adverse events (AEs)
Adverse events are defined as any undesirable experience occurring to a subject during the study, whether or not considered related to the investigational product. All adverse events reported spontaneously by the subject or observed by the investigator or his staff will be recorded.

Serious adverse events (SAEs)
A serious adverse event is any untoward medical occurrence or effect at any dose that: results in death; is life threatening (at the time of the event); requires hospitalisation or prolongation of existing inpatients' hospitalisation; results in persistent or significant disability or incapacity; is a congenital anomaly or birth defect; Any other important medical event that may not result in death, be life threatening, or require hospitalization, may be considered a serious adverse experience when, based upon appropriate medical judgement, the event may jeopardize the subject or may require an intervention to prevent one of the outcomes listed above.
The investigator will report the SAEs through the web portal ToetsingOnline to the accredited METC that approved the protocol, within 15 days after the investigator has first knowledge of the serious adverse events. SAEs that result in death or are life threatening should be reported expedited. The expedited reporting will occur not later than 7 days after the responsible investigator has first knowledge of the adverse event. This is for a preliminary report with another 8 days for completion of the report.

Suspected unexpected serious adverse reactions (SUSARs)
Adverse reactions are all untoward and unintended responses to an investigational product related to any dose administered.
Unexpected adverse reactions are SUSARs if the following three conditions are met: 1. the event must be serious (see chapter 9.2.2); 2. there must be a certain degree of probability that the event is a harmful and an undesirable reaction to the medicinal product under investigation, regardless of the administered dose; 3. the adverse reaction must be unexpected, that is to say, the nature and severity of the adverse reaction are not in agreement with the product information as recorded in: -Summary of Product Characteristics (SPC) for an authorised medicinal product; -Investigator's Brochure for an unauthorised medicinal product.
The investigator will report expedited the following SUSARs through the web portal The expedited reporting of SUSARs through the web portal ToetsingOnline is sufficient as notification to the competent authority.
The investigator will report expedited all SUSARs to the competent authorities in other Member States, according to the requirements of the Member States. The expedited reporting will occur not later than 15 days after the investigator has first knowledge of the adverse reactions. For fatal or life threatening cases the term will be maximal 7 days for a preliminary report with another 8 days for completion of the report.
Since the treating physician is not blinded, there is no need to break the code in case of an SUSAR.

Annual safety report
In addition to the expedited reporting of SUSARs, the investigator will submit, once a year throughout the clinical trial, a safety report to the accredited METC, competent authority, and competent authorities of the concerned Member States.
This safety report consists of: − a list of all suspected (unexpected or expected) serious adverse reactions, along with an aggregated summary table of all reported serious adverse reactions, ordered by organ system, per study; − a report concerning the safety of the subjects, consisting of a complete safety analysis and an evaluation of the balance between the efficacy and the harmfulness of the medicine under investigation.

Follow-up of adverse events
All AEs will be followed until they have abated, or until a stable situation has been reached. Depending on the event, follow up may require additional tests or medical procedures as indicated, and/or referral to the general physician or a medical specialist.
SAEs need to be reported till end of study within the Netherlands, as defined in the protocol.

Data Safety Monitoring Board (DSMB)
No DSMB is established, since this is a daily practice study. In case of life-threatening diseases usually the implementation of a DSMB is indicated from an ethical point of view.
But hand eczema is a non-critical indication. Frequent laboratory assessments will reduce the possibility of serious adverse events to a minimum. The patient population in this clinical trial exists of legal competent adults and the study drugs alitretinoin and azathioprine are well-investigated, well-characterized drugs.

STATISTICAL ANALYSIS
This trial is designed to demonstrate a superior response to azathioprine compared to alitretinoin in the treatment of severe chronic non-hyperkeratotic hand eczema. Response to treatment is defined as an improvement of ≥ 2 steps on the Physician Global Assessment (PGA) developed by Ruzicka et al (8,9) at 24 weeks of treatment.
All analysis will be based on the intention-to-treat principle to guard against attrition bias.
Subjects might not only withdraw because the study drug works insufficient, they might also withdraw when their hand eczema is cured.
Missing values will be handled in a way that is dependent on assumptions about the missing data. If the extent and pattern of missing data is known (e.g. missing at random (MAR), missing completely at random (MCAR), missing not at random (MNAR)), an analysis will be chosen that is valid under a plausible assumption about the missing data (probably mixed models). This is according to a strategy proposed by White et al. (28)

Severity of hand eczema
Between-group difference in response to treatment between baseline and 24 weeks of treatment. For comparison of proportions in the alitretinoin and azathioprine group the chi-square test, or Fisher's exact test if appropriate, will be used.

Severity of hand eczema
• Between-group difference in mean change in PGA between baseline and 12 and 24 weeks. For comparison of mean change between the alitretinoin and azathioprine group at week 12 and 24, the Student's t-test or Mann-Whitney Utest will be used, depending on distribution of data.
• Between-group difference in response to treatment between baseline and 12 weeks of treatment. For comparison of proportions in the alitretinoin and azathioprine group the chi-square test will be used.
• Between-group difference in mean change between baseline and week 4, 8, 12 and 24, assessed by the Hand Eczema Severity Index (HECSI) score. This will be described graphically. For comparison of mean change between the alitretinoin and azathioprine group at week 12 and 24, the Student's t-test or Mann-Whitney U-test will be used, depending on distribution of data.
• Between-group difference in time to response (time to first PGA improvement of ≥ 2 steps). This will be analysed using a Kaplan-Meier product limit estimator.
For comparison between the alitretinoin and azathioprine group the Log-rank test will be used.

Patient rated outcome measures (PROMs):
Quality of life

Safety and tolerability
• Adverse events in both groups will be registered.

Cost-utility and cost-effectiveness
• For each group (alitretinoin and cyclosporine), the mean EQ-5D scores overall and of each dimension will be reported. Results from the descriptive system of the EQ-5D-5L will be converted to a utility index value, a population based (social) value specific for the Netherlands. With this value, Dutch utility values will be calculated in order to determine the quality adjusted life years (QALYs) over the study period. Mean values of the EQ-VAS will be reported with a 95% confidence interval. For comparison of means, the Student's t-test or Mann-Whitney U-test will be used, depending on distribution of data.
• The incremental cost-effectiveness ratio (ICER) will be calculated and reported as €/QALY. Effect will be assessed with the PGA. Costs will be assessed as described in paragraph 8.2.
• A regression model will be used to estimate the association between QALYs and the PGA.

Other study parameters
Continuous data with normal distribution will be presented as mean ± SD. Variables with a skewed distribution will be presented as median and range. Categorical variables will be expressed as frequencies and percentages. Confidence intervals (CIs) are expressed to a level of 95%.
To test for normal distribution of data, the data will be visually inspected, a Q-Q plot will be constructed and in case of doubt a test for normality will be used (e.g. Shapiro-Wilk test/Kolmogorov-Smirnov test).
For comparing normally distributed continuous data, we will use the Student's t-test. In variables with non-homogeneity of variance, the Welch test will be used.
For comparing non-normally distributed continuous data, the Mann-Whitney U test will be used.
For comparing categorical data the chi-square test will be used and the Fisher's exact test will be used in variables containing low numbers.
All P-values are two-sided and a P-value < 0,05 is considered statistically significant.
Statistical analysis will be performed using SPSS 22 (SPSS Inc, Chicago, IL, USA).

Interim analysis
Not applicable.

Regulation statement
This study will be conducted according to the principles of the Declaration of Helsinki

Recruitment and consent
It is the responsibility of the investigator, or a person designated by the investigator, to obtain written informed consent from each individual participating in this study after adequate explanation of the aims, methods, objectives and potential hazards of the study. It will also be explained to patients that they are completely free to refuse to enter the study or to withdraw from it at any time for any reason. Patients will be given a period of one week to consider participation before they are asked to sign the informed consent form.
If new safety information results in significant changes in the risk/benefit assessment, the consent form will be reviewed and updated if necessary. All patients (including those already being treated) will be informed of the new information, given a copy of the revised form and asked to give their consent to continue in the study.
The patient information letter and informed consent form are attached as separate documents.

Objection by minors or incapacitated subjects
Not applicable.

Benefits and risks assessment, group relatedness
In this trial both groups are treated with a drug, known to be beneficial to hand eczema in a considerable amount of patients. So the intended benefit of both study drugs is to reduce the severity of hand eczema.
We hypothesize that azathioprine has a superior efficacy compared to alitretinoin (the registered treatment for hand eczema in The Netherlands) in severe chronic nonhyperkeratotic hand eczema. If this hypothesis is confirmed, there could be a practical, as well as a financial implication. Practically, more responding patients to azathioprine leads to a greater beneficial effect on hand eczema in this patient group. Financially, azathioprine is a lot less expensive than alitretinoin. If azathioprine shows superior efficacy in non-hyperkeratotic hand eczema, this could lead to an official registration. This, in turn, could mean a decrease in financial burden for the treatment of all nonhyperkeratotic hand eczema patients in the population.
A risk assessment is carried out in chapter 13.

Compensation for injury
The sponsor/investigator has a liability insurance which is in accordance with article 7, subsection 6 of the WMO.
The sponsor (also) has an insurance which is in accordance with the legal requirements The insurance applies to the damage that becomes apparent during the study or within 4 years after the end of the study.

Incentives
Not applicable.

Handling and storage of data and documents
Data will be handled confidentially. Data derived from the questionnaires and other paper source documents will be coded using sequential administration numbers. A subject identification code list is used to link the data to the subjects. The code is not based on the patient initials and birth-date. The code will be safeguarded by the principal investigator, dr. M.L.A. Schuttelaar. The documents will be stored in a locked room.
The digital source data will be saved in subsections of the subjects medical file. These data will be accessible to the principal investigator and the investigator, and also to the treating physician if this is not the investigator. Data will not be accessed by the blinded efficacy assessors.
All data will be recorded in electronic Case Report Forms in Utopia, software for Electronic Data Capture (, developed by the Trial Coordination Center, linked to the University Medical Center Groningen. The eCRFs will only be accessible with the username and password of the responsible investigator.
Data will be saved for 15 years after completion of this study.
All the data will be saved in accordance with the Dutch Personal Data Protection Act.

Monitoring and Quality Assurance
A monitor of the University Medical Center Groningen will carry out monitoring of this study. The monitor will get read-only access to the digital and paper documents of participants. Goal of this monitoring is to review if: the rights and wellbeing of subjects are being protected the reported data is right and fully reproducible the execution of the study is in accordance with this protocol and relevant legal requirements.

Amendments
A 'substantial amendment' is defined as an amendment to the terms of the METC application, or to the protocol or any other supporting documentation, that is likely to affect to a significant degree: the safety or physical or mental integrity of the subjects of the trial; the scientific value of the trial; the conduct or management of the trial; or the quality or safety of any intervention used in the trial.
All substantial amendments will be notified to the METC and to the competent authority.
Non-substantial amendments will not be notified to the accredited METC and the competent authority, but will be recorded and filed by the investigator.

Annual progress report
The investigator will submit a summary of the progress of the trial to the accredited METC once a year. Information will be provided on the date of inclusion of the first subject, numbers of subjects included and numbers of subjects that have completed the trial, serious adverse events/ serious adverse reactions, other problems, and amendments.

End of study report
The investigator will notify the accredited METC and the competent authority of the end of the study within a period of 90 days. The end of the study is defined as the last patient's last visit.
In case the study is ended prematurely, the investigator will notify the accredited METC and the competent authority within 15 days, including the reasons for the premature termination.
Within one year after the end of the study, the investigator will submit a final study report with the results of the study, including any publications/abstracts of the study, to the accredited METC and the Competent Authority.

Public disclosure and publication policy
This study is designed honoring the CCMO statement on publication policy.
This is an investigator initiated study. The results of the study will be made public unreservedly; they will be offered for publication in a peer reviewed journal. In a publication all data will be handled anonymously.
The study will be registered in a public trail register before the inclusion of the first patient.

STRUCTURED RISK ANALYSIS
13.1 Potential issues of concern Both alitretinoin and azathioprine are drugs that are routinely used in medicine and particularly in the treatment of hand eczema in our department. The information asked in this chapter is described in detail in the SPC text documents of the study drugs. (10,18) These texts are used as important sources on which this protocol is based.
a. Level of knowledge about mechanism of action For this section the reader is referred to paragraph 6.1 and the SPC text of alitretinoin and azathioprine. (10,18) b. Previous exposure of human beings with the test product(s) and/or products with a similar biological mechanism For this section the reader is referred to paragraph 6.3 and the SPC text of alitretinoin and azathioprine. (10,18) c. Can the primary or secondary mechanism be induced in animals and/or in ex-vivo human cell material?
For this section the reader is referred to paragraph 6.2 and the SPC text of alitretinoin and azathioprine. (10,18) d. Selectivity of the mechanism to target tissue in animals and/or human beings For this section the reader is referred to paragraph 6.1 and the SPC text of alitretinoin and azathioprine. (10,18) e. Analysis of potential effect Alitretinoin (from SPC text) (10) The recommended dose range for alitretinoin is 10 mg -30 mg once daily.
The recommended start dose for alitretinoin is 30 mg once daily. A dose reduction to 10 mg once daily may be considered in patients with unacceptable adverse reactions to the higher dose. In studies investigating 10 mg and 30 mg daily doses, both doses resulted in clearing of the disease. The 30 mg dose provided a more rapid response and a higher response rate. The 10 mg daily dose was associated with fewer adverse events.
A treatment course of alitretinoin may be given for 12 to 24 weeks depending on response. Discontinuation of therapy should be considered for patients who still have severe disease after the initial 12 weeks of treatment. In the event of relapse, patients may benefit from further treatment courses of alitretinoin. The capsules should be taken with a meal once daily.
Alitretinoin should not be prescribed if the patient's eczema can be adequately controlled by standard measures, including skin protection, avoidance of allergens and irritants, and treatment with potent topical corticosteroids.
Risks of alitretinoin treatment are described in paragraph 6.4.
Azathioprine (from SPC text) (18) In general, the starting dosage is 1-3mg/kg/body weight/day and should be adjusted according to the clinical response (which may not be evident for weeks or months) and haematological tolerance.
The maintenance dosage required may range from less than 1mg/kg body weight/day to 3mg/kg/body weight/day depending on the clinical condition being treated and the individual patient response including haematological tolerance.
Risks of azathioprine treatment are described in paragraph 6.4 and 6.5 (regarding TPMT measurement).

Absorption
The absorption of alitretinoin from the gastro-intestinal tract is variable and doseproportional over the therapeutic range from 10-30 mg. The absolute bioavailability of alitretinoin has not been determined. When alitretinoin is taken with food, the systemic exposure is enhanced by a factor of 4 and the variability of exposure is decreased.
Therefore, alitretinoin should be taken with a meal.

Distribution
Alitretinoin strongly binds to plasma proteins. The volume of distribution of alitretinoin in man has not been determined, but animal studies indicate a volume of distribution greater than the extracellular volume.

Metabolism
Alitretinoin is metabolized by oxidation in the liver by CYP3A4 isoenzymes into 4-oxoalitretinoin. Both compounds undergo isomerization into all-trans retinoic acid and 4-oxoall-trans retinoic acid. After oral administration, the contribution of the metabolites in plasma to the systemic exposure of alitretinoin is approximately 35% to 80% for 4-oxoalitretinoin. The major metabolite 4-oxo-alitretinoin is further glucuronidated and eliminated in urine. Alitretinoin is degraded similarly to vitamin A by sequential cleavage of the carbon-side chain. During a 12-to 24-week treatment period with 10 or 30 mg dose, the exposure to alitretinoin remained stable.

Elimination
Alitretinoin is an endogenous retinoid. Alitretinoin concentrations return to normal range within 1 to 3 days treatment cessation. Excretion of radio-labelled alitretinoin was complete with approximately 94% of the dose recovered. Radio-labelled material was eliminated mainly in urine and a smaller fraction (approx. 30%) in faeces. The most abundant excretion compound is the glucuronide of 4-oxo-alitretinoin amounting to 6.5% of the dose in urine. Elimination half-life of unchanged alitretinoin ranges between 2 to 10 hours. Alitretinoin and its 4-oxo-metabolite do not accumulate.

Pharmacokinetic in special populations
In a pharmacokinetic study in patients, gender, weight and age did not affect the pharmacokinetics of alitretinoin. The pharmacokinetics of alitretinoin in CHE patients was similar to that in healthy volunteers. Alitretinoin kinetics has not been studied in patients with hepatic or with severe renal insufficiency or in patients below 18 years.

Azathioprine (from SPC text) (18)
Azathioprine is well absorbed following oral administration. Peak plasma concentrations are reached 1-2 hours after taking a dose. Azathioprine is distributed rapidly throughout the body. The plasma half life is 3-5 hours. Only 30% of the medicinal product binds to plasma proteins. 12.5% enter the cerebrospinal fluid.
Azathioprine is extensively metabolised to 6-thioinosinic acid and methyl mercaptopurineribonucleotide, which, in part, are responsible for the effect of the medicinal product.
The effect in-vivo is complicated by the action of methyl-nitroimidazole, which is also found.
Up to 50% of a dose is excreted in urine during the first 24 hours after administration, with approximately 10% as unchanged substance. Only 12.6% of the dose is excreted during 48 hours with the faeces. There is no evidence for enterohepatic circulation.
A lowered dosage for patients with reduced renal function may be necessary, probably as a result of reduced elimination of the active metabolites of azathioprine.
Also in patients with hepatic impairment the metabolism of azathioprine is altered.
Conversion into the active form is reduced, and especially the breakdown to eliminable metabolites is diminished.
Mercaptopurine, a metabolite of azathioprine, has been identified in the colostrum and breast-milk of women receiving azathioprine treatment. In general, otherwise healthy subject with severe, chronic non-hyperkeratotic hand eczema will be included. Woman with child bearing potential will only be included in the study if they agree to use proper contraceptive methods during study period and up to two months following end of study.
h. Interaction with other products For this section the reader is referred to paragraph 5.2 and the SPC text of alitretinoin and azathioprine. (10,18) i. Predictability of effect No biomarkers for effect have been determined.
j. Can effects be managed?

Alitretinoin (from SPC text) (10)
Alitretinoin is a derivative of vitamin A. Alitretinoin has been administered in oncological clinical studies at dosages of more than 10-times of the therapeutic dosage given for chronic hand eczema. The adverse effects observed were consistent with retinoid toxicity, and included severe headache, diarrhoea, facial flushing, hypertriglyceridemia. These effects were reversible.
It can be concluded that the (reversible) effects can be properly managed.
Azathioprine (from SPC text) (18) Symptoms: In the event of overdose the most likely effect is bone marrow suppression, reaching its maximum mostly 9-19 days after dosing. The principal signs of bone marrow suppression are ulceration of the throat, fever and infections. Furthermore, bruising, bleeding and fatigue may occur. A single large dose of azathioprine is less likely to have a toxic effect than a chronic minor overdosage (e.g. on prescription). Although improvement may be delayed, it usually occurs from the twelfth day after overdose, provided that the patient has not taken a high dose in the meantime.
Protocol ID 52232 Alitretinoin vs azathioprine in severe non-hyperkeratotic hand eczema There is no specific antidote for azathioprine. In the event of overdose, blood count and hepatic function in particular should be monitored. Azathioprine is known to be dialysable and in severe cases dialysis may be used.
It can be concluded that management of severe effects, if needed, could be radical but possible.

Synthesis
The overall risks for patients participating in this study are acceptable because of the tight inclusion and exclusion criteria (ensuring a relatively healthy study population), combined with regular laboratory assessments to enhance safety monitoring by the investigator or treating physician. Furthermore, prior experience with both study drugs in daily practice has improved our capability to manage risks.
A specific measure that will be taken to reduce risks is the TPMT measurement. This will be carried out in the azathioprine group to exclude patients with a low or absent TPMT activity, to prevent certain life-threatening myelumsuppression to occur in these patients.
In general, adult subjects with severe chronic non-hyperkeratotic hand eczema will be included, who are otherwise healthy. The remaining risk is therefore small, does not differ considerably from regular daily practice, and is therefore deemed acceptable for study subjects.